Neuroscience Research Australia and the University of New South Wales, Sydney, 2031, Australia,
Neurosci Bull. 2013 Oct;29(5):649-54. doi: 10.1007/s12264-013-1379-y. Epub 2013 Aug 30.
Both fresh-frozen and formalin-fixed, paraffin-embedded (FFPE) human brain tissues are invaluable resources for molecular genetic studies of central nervous system diseases, especially neurodegenerative disorders. To identify the optimal method for DNA extraction from human brain tissue, we compared methods on differently-processed tissues. Fragments of LRRK2 and MAPT (257 bp and 483 bp/245 bp) were amplified for evaluation. We found that for FFPE samples, the success rate of DNA extraction was greater when using a commercial kit than a laboratory-based method (successful DNA extraction from 76% versus 33% of samples). PCR amplicon size and storage period were key factors influencing the success rate of DNA extraction from FFPE samples. In the fresh-frozen samples, the DNA extraction success rate was 100% using either a commercial kit (QIAamp DNA Micro) or a laboratory-based method (sample boiling in 0.1 mol/L NaOH, followed by proteinase K digestion, and then DNA extraction using Chelex-100) regardless of PCR amplicon length or tissue storage time. Although the present results demonstrate that PCR-amplifiable genomic DNA can be extracted from both fresh-frozen and FFPE samples, fresh brain tissue is recommended for DNA extraction in future neuropathological studies.
新鲜冷冻和福尔马林固定、石蜡包埋（FFPE）的人脑组织是研究中枢神经系统疾病（尤其是神经退行性疾病）分子遗传学的宝贵资源。为了确定从人脑组织中提取 DNA 的最佳方法，我们比较了不同处理组织的方法。评估了 LRRK2 和 MAPT（257 bp 和 483 bp/245 bp）片段的扩增情况。我们发现，对于 FFPE 样本，使用商业试剂盒比实验室方法的 DNA 提取成功率更高（成功提取 DNA 的样本比例分别为 76%和 33%）。PCR 扩增子大小和储存时间是影响 FFPE 样本 DNA 提取成功率的关键因素。在新鲜冷冻样本中，使用商业试剂盒（QIAamp DNA Micro）或实验室方法（样本在 0.1 mol/L NaOH 中煮沸，然后用蛋白酶 K 消化，再用 Chelex-100 提取 DNA），无论 PCR 扩增子长度或组织储存时间如何，DNA 提取成功率均为 100%。尽管目前的结果表明，可以从新鲜冷冻和 FFPE 样本中提取可用于 PCR 的基因组 DNA，但在未来的神经病理学研究中，建议使用新鲜脑组织进行 DNA 提取。
